Local thermodynamics govern formation and dissolution ofCaenorhabditiselegans P granule condensates

Abstract

Membraneless compartments, also known as condensates, provide chemically distinct environments and thus spatially organize the cell. A well-studied example of condensates is P granules in the roundworm Caenorhabditis elegans that play an important role in the development of the germline. P granules are RNA-rich protein condensates that share the key properties of liquid droplets such as a spherical shape, the ability to fuse, and fast diffusion of their molecular components. An outstanding question is to what extent phase separation at thermodynamic equilibrium is appropriate to describe the formation of condensates in an active cellular environment. To address this question, we investigate the response of P granule condensates in living cells to temperature changes. We observe that P granules dissolve upon increasing the temperature and recondense upon lowering the temperature in a reversible manner. Strikingly, this temperature response can be captured by in vivo phase diagrams that are well described by a Flory–Huggins model at thermodynamic equilibrium. This finding is surprising due to active processes in a living cell. To address the impact of such active processes on intracellular phase separation, we discuss temperature heterogeneities. We show that, for typical estimates of the density of active processes, temperature represents a well-defined variable and that mesoscopic volume elements are at local thermodynamic equilibrium. Our findings provide strong evidence that P granule assembly and disassembly are governed by phase separation based on local thermal equilibria where the nonequilibrium nature of the cytoplasm is manifested on larger scales.

Sequence- and chemical specificity define the functional landscape of intrinsically disordered regions

“…If this is correct, phase separation would be an unavoidable but perhaps relatively benign consequence of these multivalent interactions ( 71, 91 ). We emphasize that there are also many examples in which intracellular phase transitions offer compelling functional advantages ( 74, 87, 92 – 97 ). Delineating functionally important phase transitions from situations in which it is an unavoidable outcome of multivalent proteins is a major and open challenge for the field of cell biology.…”

Phase separating RNA binding proteins form heterogeneous distributions of clusters in subsaturated solutions

“…For a purely phase separationbased process described by classical Flory-Huggins style theories (10, 11), subsaturated solutions should feature dispersed monomers and very few small clusters, if any, at any given time. It is worth noting that these simple theories have been the mainstay for quantitative descriptions and analysis of phase transitions driven by multivalent proteins and nucleic acids in vitro and in cells (12–15, 21, 42, 57–62). Surprisingly, we find that subsaturated solutions do not conform to expectations based on systems that are characterized by a single energy scale such as the Flory χ -parameter.…”

LIN-35 is necessary in both the soma and germline for preserving fertility in Caenorhabditis elegans under moderate temperature stress

“…We found that for all strains at 20°C, the vast majority of L1 PGCs showed PGL-1 with a normal pattern of localization in puncta that are relatively evenly distributed around the nucleus of the two PGCs (Fig 6a-b). However, we found that for all strains, except lin-35(n745); pie-1p::lin-35, there was a significant increase in the number of L1s that had PGL-1 mislocalized within PGCs at 26°C compared to the same Previous work has also shown that PGL-1 can become diffuse in adult germlines subjected to short exposures of 29°C, specifically in the distal part of the germline near the mitotic zone and in oocytes [33]. We therefore looked to see if we could also see changes to PGL-1 localization in adult germlines of hermaphrodites that spent their entire life at 26°C.…”

“…Previous work has also shown that PGL-1 can become diffuse in adult germlines subjected to short exposures of 29°C, specifically in the distal part of the germline near the mitotic zone and in oocytes [33]. We therefore looked to see if we could also see changes to PGL-1 localization in adult germlines of hermaphrodites that spent their entire life at 26°C.…”